The largest prison outbreak of TB in Western Europe investigated using whole-genome sequencing.

BACKGROUND: In March 2011, the Department of Public Health East in Ireland were notified of two cases of TB in two prisoners sharing a cell. We define the resulting outbreak and highlight the role of public health and laboratory-based molecular epidemiology in mapping and control of a prison outbreak.METHODS: Cases were identified through clinical presentation, contact tracing, case-finding exercise or enhanced laboratory surveillance. Mycobacterium tuberculosis isolates were genotyped and underwent whole-genome sequencing (WGS).RESULTS: Of the 34 cases of TB linked to the outbreak, 27 were prisoners (79%), 4 prison officers (12%) and 3 community cases (9%). M. tuberculosis was isolated from 31 cases (culture positivity: 91%). A maximum of six single-nucleotide polymorphisms separated the isolates, with 22 being identical, suggestive of a highly infectious 'super-spreader´ within the prison. Isolates belonged to the Beijing sub-lineage, and were susceptible to first-line anti-TB agents. A case-finding exercise incidentally detected a prisoner with multidrug-resistant TB. Of the 143 prison officers screened, 52% had latent TB infection. Litigation costs exceeded five million euros.CONCLUSION: This constitutes the largest prison outbreak of TB in Western Europe investigated using WGS. A robust prison entry TB screening and education programme is required to effect better TB control, and prevent future outbreaks and attendant litigation.

Directions of motion after-effects induced by gratings and plaids.

In three experiments the direction of motion after-effect (MAE) is measured following adaptation to two gratings moving in different directions presented in alternation (component-induced MAEs: CMAEs), and to moving plaid patterns composed of superimposed pairs of these gratings (plaid-induced MAEs; PMAEs). These MAEs are compared to: (i) the vector sum direction of the component gratings; (ii) the IOC-predicted direction of the plaids; and (iii) the perceived direction of the plaids as reported by observers. Contrary to previous findings (Burke D, Wenderoth P. Vis Res 1993;33:351-9), directions of PMAEs are shown to approximate the vector sum direction of the components, whereas directions of CMAEs are shown to approximate the mean (unweighted) direction of the components. This difference is attributed to the activity, and adaptation, of an additional population of neurones whose stimulus), or a counterphase moving plaid (a combined Fourier and non-Fourier stimulus), rules out the possibility that the discrepancy between PMAE direction and actual plaid direction is due to the use of test stimuli that do not adequately reflect adaptation by the Fourier and non-Fourier components of the adapting plaids (HR, Ferrera VP, Yo C. Vis Neurosci 1992;9:79-97). Various explanations of this paradoxical result are discussed, including: (i) that MAEs produced by Fourier components out-weigh (and possibly even mask) MAEs produced by non-Fourier plaid components; (ii) PMAEs are influenced by adaptation of a population of component-selective neurones that do not contribute to plaid perception; and, (iii) PMAEs are influenced by component-specific adaptation effects that are weighted according to relative component sensitivity, rather than relative component speed (Pantle A. Vis Res 14;1974:1229-36). We review psychophysical and neurophysiological evidence consistent with these explanations.

Iron-responsive element-binding protein in hemochromatosis liver and intestine.

Iron-responsive element-binding protein (IRE-BP) activity was studied in liver and intestinal samples of hemochromatosis and control patients using a short 32P-IRE-RNA probe on "retardation" nondenaturing polyacrylamide gels. IRE-BP activity was assessed in liver biopsy specimens in 36 patients--16 hemochromatosis homozygotes, 4 hemochromatosis heterozygotes, 6 patients with secondary iron overload, and 10 control patients with normal hepatic iron concentrations. Intestinal IRE-BP activity was assessed in 14 hemochromatosis homozygotes and 16 normal subjects. Endogenous IRE-BP activity was determined from 32P retarded on the gel, and total IRE-BP activity was assessed after reducing tissue samples with 2-mercaptoethanol. Hepatic endogenous IRE-BP activity was inversely related to hepatic iron concentration (r = .59, P < .0002). Mean hepatic endogenous IRE-BP activity in the hemochromatosis homozygotes, 0.25 +/- 0.04 pmol/mg protein, was significantly decreased compared with values in the normal controls, 0.45 +/- 0.06 pmol/mg protein, P < .05. Hepatic total IRE-BP was also significantly decreased in the hemochromatosis patients by gel retardation assay and Western blotting with anti-IRE-BP antibody. Intestinal endogenous IRE-BP activity, total IRE-BP activity, and iron concentration did not significantly differ between hemochromatosis patients and normal control subjects. This suggests that both endogenous IRE-BP activity and the total amount of the protein are downregulated in the liver by tissue iron. Intestinal IRE-BP activity that regulates intestinal transferrin receptor expression is normal in hemochromatosis and appropriate for the intracellular iron concentration.

Monocyte membrane ferritin in hemochromatosis.

To further evaluate a possible abnormality in the reticuloendothelial cells in hemochromatosis, the binding of a monoclonal anti-human liver ferritin antibody to monocytes was studied in 19 patients with hemochromatosis, 8 patients with secondary iron overload, 1 patient with hyperferritinemia without iron overload, and 15 normal volunteers. Binding of the antibody to the monocytes was analyzed using a fluorescence-activated cell sorter (FACS). Binding of the anti-ferritin antibody to monocytes was demonstrated in 34.7 +/- 4.5% (mean +/- standard error) of the monocytes in untreated hemochromatosis patients (mean serum ferritin = 2294 +/- 415 micrograms/L), 6.75 +/- 2.03% in treated hemochromatosis patients (mean serum ferritin = 263 +/- 85 micrograms/L), 12.3 +/- 2.7% of the monocytes in the secondary iron overload patients (mean serum ferritin = 2476 +/- 867 micrograms/L), 4.1% in the patient with hyperferritinemia (serum ferritin = 1192) and 4.1 +/- 0.5% of the monocytes in the normal volunteers (mean serum ferritin = 55.2 +/- 11.9 micrograms/L). % binding of anti-ferritin antibody was significantly greater in hemochromatosis patients compared to patients with secondary iron overload (p less than 0.05) despite a comparable degree of iron overload in the secondary iron overload group. The addition of exogenous human ferritin to samples from treated hemochromatosis patients and normal volunteers did not significantly increase the % of monocytes binding anti-ferritin antibody. These results suggest that monocytes from iron-loaded hemochromatosis patients express increased surface ferritin which may represent release of ferritin and a metabolic defect characteristic of hemochromatosis.

Mucosal iron in the control of iron absorption in a rat intestinal transplant model.

Isogeneic intestinal transplantation of iron-loaded and iron-deficient intestine into iron-deficient rats was performed in 20 Lewis rats to isolate the effect of intestinal mucosal iron on iron absorption. Rats were iron loaded with three weekly IM injections of 50 mg of iron dextran and were rendered iron deficient with an iron-deficient diet for 3 weeks. Iron status was assessed by hepatic and gut mucosal iron determination. Uptake and transfer of 59Fe-ascorbate was measured in an isolated perfused segment of transplanted intestine 48 hours after transplantation. The mean rate of uptake of 59Fe from an iron-loaded intestine (mean mucosal iron concentration, 7.97 +/- 2.02 mumol/g) was 431 +/- 27 nmol/30 min, and from an iron-deficient intestine (mean mucosal iron concentration, 1.35 +/- .84 mumol/g), 743 +/- 222 nmol/30 min (P less than 0.001). The mean transfer of 59Fe from the mucosal cell to the body through an iron-loaded intestine was 63 +/- 22 nmol/30 min, and through an iron-deficient intestine was 86 +/- 32 nmol/30 min (P less than 0.05). These results suggest that the gut mucosal iron concentration regulates the uptake and transfer of iron in the intestine.

Abnormalities in iron metabolism in multiple sclerosis.

High iron concentrations have been reported in the brains of multiple sclerosis victims. To determine if there are abnormalities in general iron metabolism indicative of iron overload in MS, measurements of transferrin saturation, serum ferritin and red cell ferritin in 31 female and 18 male patients were compared to the results in 49 age- and sex-matched healthy controls. Compared to controls, mean serum ferritin in MS was high, whereas transferrin saturation and red cell ferritin were similar. High values in one or more individual test results were observed in eleven MS patients. They were prevalent in patients who required bilateral assistance to walk or were confined to a chair, and appeared to be related to the severity of the disease. An investigation was made into the relationship of the high serum ferritin values in MS to the HLA-A3 histocompatibility antigen, a marker of the hemochromatosis gene which is prevalent in MS. A statistically significant interaction was not found between serum ferritin and the presence of HLA-A3.

Ferritin release by mononuclear cells in hereditary hemochromatosis.

An anomaly of the iron-loading disorder hereditary hemochromatosis is that bone marrow iron stores remain low until later stages of the disease. The possibility that this may be related to a disorder of reticuloendothelial ferritin metabolism was examined by studying ferritin release from mononuclear cells. Ferritin release was measured in peripheral blood mononuclear cells from four patients with hemochromatosis who had not received treatment, from six patients with hemochromatosis who had received treatment, and from 10 age- and gender-matched controls by using a modified hemolytic plaque assay. Ferritin release from the hemochromatotic cells was enhanced when compared with that of controls, and added iron stimulated ferritin release to a comparable degree in both groups. Enhanced ferritin release above matched control values was found both in cells from patients with hemochromatosis with partial phlebotomy who had high serum ferritin values and in cells from patients with hemochromatosis with full phlebotomy who had normal serum ferritin values. The increased ferritin release observed in these studies may signify abnormal reticuloendothelial iron metabolism in hemochromatosis.

Mechanisms and regulation of intestinal uptake and transfer of iron.

Despite much research over the past fifty years, the precise details of intestinal iron absorption remain unclear. The lack of understanding extends both to the specific biochemical mechanisms of transport as well as the means by which these are regulated. Iron in several dietary forms must be digested and processed in the intestinal lumen, taken up across the mucosal brush border membrane, transported through the intestinal absorptive cell, accomplish a second transmembrane passage across the basolateral membrane and, finally, be transported to sites of iron metabolism and storage within the body. Recent findings in several of these areas are reviewed. Evidence is presented to support the involvement, or non-involvement, of several intestinal iron-binding components in iron transport. In the future, the application of molecular biology to the investigation of intestinal iron metabolism will undoubtedly increase understanding of absorption mechanisms and their regulation.

Usefulness of erythrocyte ferritin analysis in hereditary hemochromatosis.

A study was carried out to determine the usefulness of erythrocyte ferritin analysis in identifying homozygotes and heterozygotes in families affected with hereditary hemochromatosis, an autosomal recessive disorder. To select the subjects the genotypes of 60 people from 26 affected families were determined by HLA-A and HLA-B haplotyping. In addition, data for 12 homozygotes for whom erythrocyte ferritin values were available from the literature were included. Likelihood analysis was used to evaluate the diagnostic value of erythrocyte ferritin analysis alone and in combination with serum ferritin testing. An erythrocyte ferritin value of 150 ag/cell or higher combined with a serum ferritin level above the 90th percentile indicated homozygosity, whereas a value of less than 150 ag/cell and a serum ferritin level at or below the 90th percentile indicated that homozygosity could be ruled out with a high degree of confidence. The probability of heterozygosity rose to 92% when the erythrocyte ferritin value was between 29 and 149 ag/cell and to 98% when this result was combined with a serum ferritin level at or below the 90th percentile. Erythrocyte ferritin analysis in combination with serum ferritin testing is useful for identifying homozygotes and a proportion of heterozygotes in families affected with hemochromatosis.

Transferrin receptors in the human gastrointestinal tract. Relationship to body iron stores.

Fluorescently labeled antibodies were used to identify transferrin receptors and mucosal transferrin in human gastrointestinal biopsy sections. Transferrin receptors were evident in the villous epithelium and the crypt areas of duodenum, ileum, and colon, predominantly in the basal-lateral area. In 7 subjects with low iron stores, the intensity of duodenal villous staining for receptor, on a scale of 0-4, was 2.1 +/- 0.3 (mean +/- SD). This value was significantly higher than the value in 13 subjects with normal iron stores (1.1 +/- 0.4). In 5 patients with hereditary hemochromatosis, duodenal transferrin receptor staining was not significantly different from that in the subjects with normal iron stores. Transferrin staining was found in the apical cytoplasm of epithelial cells in the duodenum, ileum, and colon, but observer assessment was not sufficiently reproducible to make a quantitative analysis. Our results suggest that iron deficiency is accompanied by an increase in transferrin receptors in duodenal absorptive cells, and the genetic lesion in hemochromatosis does not involve an increase in transferrin receptors in the intestinal mucosa compared with subjects with normal iron stores.

Zinc absorption in inflammatory bowel disease.

Zinc absorption was measured in 29 patients with inflammatory bowel disease and a wide spectrum of disease activity to determine its relationship to disease activity, general nutritional state, and zinc status. Patients with severe disease requiring either supplementary oral or parenteral nutrition were excluded. The mean 65ZnCl2 absorption, in the patients, determined using a 65Zn and 51Cr stool-counting test, 45 +/- 17% (SD), was significantly lower than the values, 54 +/- 16%, in 30 healthy controls, P less than 0.05. Low 65ZnCl2 absorption was related to undernutrition, but not to disease activity in the absence of undernutrition or to zinc status estimated by leukocyte zinc measurements. Mean plasma zinc or leukocyte zinc concentrations in patients did not differ significantly from controls, and only two patients with moderate disease had leukocyte zinc values below the 5th percentile of normal. In another group of nine patients with inflammatory bowel disease of mild-to-moderate severity and minimal nutritional impairment, 65Zn absorption from an extrinsically labeled turkey test meal was 31 +/- 10% compared to 33 +/- 7% in 17 healthy controls, P greater than 0.1. Thus, impairment in 65ZnCl2 absorption in the patients selected for this study was only evident in undernourished persons with moderate or severe disease activity, but biochemical evidence of zinc deficiency was uncommon, and clinical features of zinc depletion were not encountered.

Zinc metabolism in patients on maintenance hemodialysis.

Recent studies have focused attention on the possible role of zinc depletion in the pathogenesis of uremic symptoms such as dysgeusia and impotence. The present studies were undertaken to evaluate the prevalence of zinc deficiency and abnormalities of zinc metabolism in patients with end-stage renal disease. A total of 43 stable chronic hemodialysis patients were screened for evidence of zinc deficiency by measurement of fasting predialysis leukocyte and plasma zinc. The results were compared with those from 30 healthy volunteers. Seventeen of these 43 patients had 65Zn absorption measured, and in 9 the rate of disappearance of 65Zn from the body was also measured. The results were compared with those obtained in 20 healthy controls. The nutritional status of these 17 patients was estimated by global nutritional assessment and calculation of the Quetelet index while 9 of 17 had dietary zinc intake calculated from a diet history. The mean plasma zinc level was lower in the hemodialysis patients (11.7 +/- 2.3 mumol/l vs. 13.3 +/- 2.9 mumol/l in controls; p less than 0.05). The mean leukocyte zinc level was 0.81 +/- 0.27 mumol/g dry weight in dialysis patients and 0.81 +/- 0.22 mumol/g in controls (p greater than 0.2). The mean 65Zn absorption in the patients was 49 +/- 14% and in controls 53 +/- 12% (p greater than 0.2). Mean turnover of body 65Zn was 0.47 +/- 0.11%/day in patients and 0.43 +/- 0.18%/day in controls (p greater than 0.1). The mean 65Zn half-life was 154 +/- 29 days in patients and 187 +/- 75 days in controls (p greater than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Zinc absorption and leukocyte zinc in alcoholic and nonalcoholic cirrhosis.

To determine if malabsorption of zinc contributes to the zinc deficiency found in cirrhosis, the absorption of an oral dose of ZnCl2, labeled with 65Zn and a nonabsorbed marker 51CrCl3, was determined from the ratio of these isotopes in a stool specimen. Average 65Zn absorption in 25 alcoholic cirrhotics, 37 +/- 17% (SD), was low compared to 55 +/- 16% in 31 healthy volunteer controls (P less than 0.01). In contrast, mean 65Zn absorption, 47 +/- 11%, in 11 nonalcoholic cirrhotics was not significantly different from the average result in healthy controls. Low 65Zn absorption was accompanied by low leukocyte zinc in a subgroup of alcoholic cirrhotics with ascites and/or ascites and encephalopathy, but not in the subgroup in which these clinical features were absent. Thus, low zinc absorption contributes to zinc deficiency in decompensated alcoholic cirrhosis. The failure to find similar abnormalities in nonalcoholic cirrhosis suggests that the long-standing consumption of alcoholic beverages contributes to the malabsorption of zinc.

Dual-isotope method for determination of human zinc absorption: the use of a test meal of turkey meat.

The percentage of 65Zn taken up (absorbed) from extrinsically labeled turkey meat was calculated from the amounts of 65Zn and a nonabsorbed 51Cr marker present in the body or in a single stool specimen after 1-2 d. 51CrCl3 proved to be a suitable marker for unabsorbed 65Zn and so the early determination of 65Zn absorption was possible. With stool counting, 65Zn absorption data from first stool samples after 1-2 d were accurate as judged by correlation with the amount of 65Zn in the body 7-10 d later (retention); results from subsequent stools gave lower absorption values due to the early excretion of some absorbed 65Zn. The dual-isotope method gave reproducible results when four successive tests of zinc absorption were carried out in a group of six subjects. The average (mean +/- SD) 65Zn absorption from turkey meals containing 31 mumol (2 mg) and 46 mumol (3 mg) of zinc was 39 +/- 8% and 29 +/- 6%, respectively, measured by stool counting; 65Zn absorption and retention correlated well in both studies. A series of different beverages was given in place of water with the turkey meal. Orange juice significantly reduced 65Zn absorption and milk also showed this tendency, but tea, whiskey, wine or beer had no significant effect on the absorption of 65Zn from the turkey meal. In groups of subjects the mean ratio of 65Zn absorption from extrinsically labeled turkey meat on two occasions (1.06) was not significantly different from that of the absorption of extrinsic to intrinsic 65Zn labels (1.16). The dual-isotope technique with either stool or body counting is suitable for the rapid determination of 65Zn absorption from extrinsically labeled turkey within 2 d.

Does the oral zinc tolerance test measure zinc absorption?

Increases in plasma zinc concentration were compared with radiozinc absorption after oral test doses. Ten healthy, fasting subjects were each given 385 mumol zinc chloride (25 mg Zn) labelled with 0.5 muCi 65ZnCl2 and a non-absorbed marker, 51CrCl3, dissolved in 100 ml of water; another 10 persons were given 354 mumol zinc chloride and 125 g of minced turkey containing 31 mumol zinc also labelled with 65Zn and 51Cr. Measurements were made of plasma zinc concentration at hourly intervals for 5 hours, radiozinc absorption by stool counting of unabsorbed radioactivity 12-36 hours later, and radiozinc retention by whole body counting at 7 days. The mean percentage of radiozinc absorbed and retained in the body from the two test meals was found to be identical (42%). In contrast the increased area under the plasma zinc curve up to 5 hours after the turkey meal, 28 +/- 9 mumol/L (mean +/- SD) was significantly less than that for zinc chloride alone, 47 +/- 15 mumol/L, p less than 0.005. Despite this difference, a good correlation was found between the area under the plasma zinc curve and 65Zn absorption in individual subjects after each meal. The discrepancy between the results of zinc absorption derived from the plasma zinc curve and 65Zn absorption for the liquid and solid test meals was most likely explained by binding of zinc to food and delayed gastric emptying of the solid meal. With a test meal of turkey meat at least this dampened the plasma appearance of zinc but did not affect its overall absorption.

Intestinal absorption of zinc: competitive interactions with iron, cobalt, and copper in mice with sex-linked anemia (sla).

Duodenal uptake and transfer of 59Fe and 65Zn and absorptive interactions between iron, zinc, cobalt, and copper were studied in sla mice and in genetically normal Swiss albino control mice. Genetically normal mice with a high iron-absorbing capacity, induced by being fed an iron-deficient diet, showed greater uptake and transfer of 65Zn in duodenum but not ileum, compared with mice with a low iron-absorbing capacity. 59Fe transfer from the duodenal mucosa to the body was lower in sla mice compared with controls and bleeding stimulated the capacity to absorb 59Fe less in sla mice relative to normal controls. In contrast, 65Zn transfer in sla was no different from controls and was not stimulated by bleeding in sla or in control mice. Iron or cobalt in a 10-fold molar excess predominantly lowered 65Zn transfer in both sla and controls, but in a study of the effect of zinc on iron transport only the uptake of 59Fe in sla mice was lowered by excess zinc in the perfusate. The effect of added copper on 65Zn transport was paradoxical; in both sla and control mice copper markedly increased 65Zn uptake relative to perfusates containing 65Zn alone, but transfer in normal mice was lowered whereas it was increased in sla animals. The interaction between zinc and iron does not appear to take place at the site of the genetic defect in sla mice. The lesion in iron transport in these mice is likely due to defective binding and transfer sites in the basolateral membrane; these sites are apparently exclusive for iron and not shared by zinc.

Effects of iron, tin, and copper on zinc absorption in humans.

Zinc absorption as measured by body retention of [65Zn]zinc chloride or a turkey test meal extrinsically labeled with 65Zn was determined in human subjects by whole body counting after 7 days. Average 65Zn absorption from zinc chloride in persons with a high iron-absorbing capacity was similar to persons with a low capacity to absorb iron. Inorganic iron, 920 mumol (51 mg), or HB iron, 480 mumol (26 mg), inhibited 65Zn absorption from 92 mumol (6 mg) of zinc chloride. When 610 mumol of iron (34 mg) was added to a turkey test meal containing 61 mumol of zinc (4 mg), 65Zn absorption was not inhibited. Tin, 306 mumol (36 mg), given with zinc chloride or turkey test meals (61 mumol, 4 mg, of Zn) significantly reduced 65Zn absorption. Copper, 79 mumol (5 mg), had no significant effect on the 65Zn absorption from 7.9 mumol (0.5 mg) of zinc chloride. In summary, the capacity to absorb iron did not influence 65Zn absorption, but both inorganic iron and heme-iron inhibited 65Zn absorption from zinc chloride. Inorganic iron had no effect, however, on 65Zn absorption from the turkey test meal. Tin in a large dose also inhibited 65Zn absorption from both zinc chloride and the turkey test meal.

A model to produce pure zinc deficiency in rats and its use to demonstrate that dietary phytate increases the excretion of endogenous zinc.

An early effect of zinc deficiency in rats is loss of appetite with consequent malnourishment. To obviate the need for pair-feeding, rats were fed liquid semipurified diets by gastric tube feeding. Rats tube fed a zinc-deficient diet at a daily rate of 90-110 g/kg thrived for 6-7 days and then rapidly became seriously ill. In contrast rats fed the deficient diet ad libitum stopped growing after 3 days but remained relatively healthy. Animals tube fed a zinc-replete diet or the deficient diet with subcutaneous injections of zinc grew normally and suffered no ill effects. Rats tube fed the deficient diet and supplemented parenterally with zinc excreted significantly more endogenous zinc into small intestinal luminal washings and into feces than unsupplemented rats. Sodium phytate, added to the tube feed in three groups of rats fed the deficient diet and supplemented daily with 0, 0.33 and 3.3 mg Zn/kg, significantly increased the zinc content of luminal washings in all three groups, increased fecal zinc excretion in two groups and lowered body zinc levels, estimated by femur zinc, in two groups. Tube feeding provides a means to study: 1) pure zinc deficiency without malnourishment of other nutrients; and 2) the excretion of endogenous zinc into the gastrointestinal tract.

Zinc absorption, intraluminal zinc and intestinal metallothionein levels in zinc-deficient and zinc-replete rodents.

Zinc absorption, endogenous luminal zinc and intestinal metallothionein were examined in rats and mice fed zinc-deficient and zinc-replete diets for 5-7 days. Small intestinal luminal washings from undosed rodents fed the replete diet contained 18- to 20-fold more zinc than those from zinc-deficient animals. Isotope dilution by this endogenous zinc could account for apparent differences between the groups in absorption of 65Zn from oral doses of 0.1-0.2 mumol 65Zn. With larger (1-2 mumol) oral doses of 65Zn in mice, no difference between the dietary groups was found, although zinc-deficient rats still absorbed more zinc than zinc-replete controls. In mice, zinc absorption from perfused duodenum, jejunum and ileum also was not increased by zinc deficiency. With oral doses, food generally lowered zinc absorption where endogenous or exogenous zinc was present in the gastrointestinal tract; food in perfusates lowered uptake more than transfer. Polarographically determined metallothionein was highest in duodenum of mice. It was lowered by feeding the deficient diet for 5 days without an increase in zinc absorption. It was concluded that zinc absorption is homeostatically increased by zinc deficiency in rats but not in mice and that intestinal metallothionein levels do not affect zinc absorption in mice.